Figure 1.
Detection of the hTAAR5 receptor protein.
Expression of the rhodopsin-tagged hTAAR5 receptor in transfected, fixed HANA3A cells was detected by the anti-rhodopsin antibody 4D2 and a secondary antibody labeled with the fluorescent dye Alexa Fluor 488 (green). Cell nuclei were stained by DAPI (blue). Left: Cells transfected with hTAAR5, right: mock-transfected control cells. Scaling bar: 20 µm.
Figure 2.
Chemical structure of various tested TMA analogs.
Only tertiary amines (1) trimethylamine and (2) dimethylethylamine can activate hTAAR5. (3) triethylamine, (4) diethylmethylamine, (5) dimethylamine, (6) methylamine, (7) trimethylphosphine, (8) cyclohexylamine, (9) N-methylpiperidine, (10) pyridine, (11) β-phenylethylamine, (12) skatole, (13) ethanolamine, (14) putrescine, (15) isobutylamine, (16) dimethylbutylamine.
Figure 3.
Human TAAR5 is selectively activated by TMA and DMEA.
Responses of hTAAR5 to 42 different amines or amine-like substances. The concentration of all tested substances was 100 µM. Responses were normalized to the response to forskolin (10 µM). Data are given as mean ± SEM of 2–10 independent experiments, each performed in duplicates. TMA and DMEA induced signals significantly differing from mock-transfected controls (***p<0.001; **p<0.01).
Figure 4.
TMA and DMEA act as agonists at the hTAAR5 receptor in a concentration-dependent manner.
Human TAAR5 responses were normalized to the response to forskolin (10 µM). EC50 = 116 µM (TMA) and EC50 = 169 µM (DMEA). Detection threshold for TMA is 1 µM (*p<0.05) and for DMEA 30 µM (*p<0.05). Error bars represent SEM.
Figure 5.
TMA and DMEA activate hTAAR5 expressed in Xenopus oocytes. A:
IBMX induced currents in oocytes expressing the reporter channel CFTR only. B: In oocytes injected with hTAAR5 cRNA application of TMA and DMEA as well as IBMX as positive control generated currents. C: Test substance induced currents normalized to the corresponding IBMX response. TMA and DMEA induced currents significantly differing from Ringer control (***p<0.001; **p<0.01). Evoked currents were measured by two-electrode voltage clamp technique at a holding potential of −70 mM. Black bars indicate time of agonist application. Error bars represent SEM.
Table 1.
SNPs in the coding regions of hTAAR genes.